Surveillance of potential pathogens and antibiotic resistance in wastewater and surface water from Boston, USA and Vellore, India using long-read metagenomic sequencing

Abstract

Environmental sampling (wastewater) could be an efficient surveillance strategy to capture global emerging trends in the spread of antibiotic resistance. Long-read DNA sequencing can resolve the genetic context of antibiotic resistance genes (ARGs) and is a promising tool for non-culture-based monitoring of antibiotic-resistant pathogens and ARGs in environmental samples, but has not been rigorously validated against conventional methods. We tested long-read sequencing using the portable Nanopore MinION for surveying pathogens, ARGs, and antibiotic-resistant pathogens in municipal wastewater, hospital wastewater, and surface water collected from Boston, USA and Vellore, India. We compared detection of enteric pathogens by assembly of long reads, with and without short-read polishing, and unassembled raw long reads for ARGs to multiplex real-time PCR. Using real-time PCR as a benchmark, long-read metagenomics was 49% sensitive and 75% specific at pathogen detection in assembled contigs, and 16% sensitive and 100% specific at detecting 28 clinically relevant resistance genes in raw long reads. Short-read polishing did not substantially improve pathogen identification or impact ARG identification in the assembled contigs, demonstrating that short-read polishing is not required, which greatly reduces costs. The high specificity of ARG detection supports portable long-read sequencing as a valuable tool to profile ARGs and antibiotic-resistant pathogens for environmental surveillance programs.

Widespread Surveillance of Emerging Contaminants in the Yamuna River Basin: Evaluation of Surface Water, Sediments, Groundwater, and Aquatic Plants

“…Antibiotics in India are available over the counter, and their consistent presence in the surface waters may lead to the emergence of resistant microbial communities. Fuhrmeister had reported the presence of antibiotic resistant genes in the surface water collected from southern part of India. Apart from surface water, these genes may also pass to the other environmental compartments including sediments; groundwater even enter human beings via drinking water sourced from surface water .…”

Simultaneous detection of pathogens and antimicrobial resistance genes with the open source, cloud-based, CZ ID pipeline

“…Lastly, to highlight the application of the CZ ID AMR module for environmental surveillance of AMR pathogens, we analyzed publicly-available short-read mNGS data from a wastewater surveillance study comparing Boston, USA to Vellore, India 29 . In this study, municipal wastewater, hospital wastewater, and surface water samples were collected from each city and underwent DNA mNGS.…”

Tracking Sources and Dissemination of Indicator Antibiotic Resistance Genes at a Watershed Scale

“…A multi-drug-resistant (fluoroquinolone; glycylcycline; tetracycline; diaminopyrimidine; nitrofuran) indicator ARG oqxA was selected based on its suitability to represent 72 correlated ARGs . There is increasing evidence of the presence of oqxA in surface water, wastewater treatment plants, hospital discharge, etc. − Results demonstrated high detection frequencies and a high abundance of oqxA , and the gene was correlated with the human MST marker and urban land-use patterns in West Run. These results provide evidence of anthropogenic activity (specifically, human waste from sewer/septic failure or illicit discharge) as the source of oqxA .…”